儿童肿瘤化疗前后胃肠道粘膜屏障功能的测定及其意义
发布时间：2018-09-08 07:21
【摘要】：目的：二胺氧化酶（Diamine oxidase，DAO）存在于哺乳动物的粘膜或绒毛上层,其中大部分存在于小肠粘膜绒毛,极少部分存在于子宫内膜绒毛中；DAO是一种具有高度活性的细胞内酶,它的活性高低与绒毛高度及粘膜细胞的核酸、蛋白合成关系密切,是反映小肠粘膜结构及功能的较为理想的指标之一。组织和血浆中DAO含量变化来自于两条途径：其一是由于坏死的肠粘膜细胞脱落入肠腔,随之引起肠粘膜DAO活性的降低；其二则是肠腔内的DAO进入肠细胞间隙淋巴管和毛细血管,引起血液中DAO水平升高。因此,在临床实验和动物实验的研究中,我们可以通过测定血液中及小肠组织中的DAO水平变化，来反映小肠的粘膜屏障功能状态,尤其是能在无创条件下测定血中DAO的活性来反映肠道损伤和修复情况。脂肪酸结合蛋白(Fatty acid binding protein，FABP)是一组低分子量(15kD左右)结合长链脂肪酸的结合蛋白，其组织特异性较强，目前研究认为它存在于哺乳动物的心肌、小肠、肝脏、脂肪组织、脑、表皮等的特定组织部位，是监测组织损伤较为新兴且价值比较高的生物学指标之一；目前脂肪酸结合蛋白的测定多应用于：慢性心力衰竭(H-FABP)，肝脏移植排斥反应(L-FABP)，非心跳供体肾的存活性选择(H-FABP、L-FABP)；专业运动员肌肉损伤的预防(S-FABP)等的研究中。肠脂肪酸结合蛋白(Intestinal fatty acid binding protein，I-FABP)作为目前已发现的FABP的九种类型之一，它的组织特异性高，因其全部来源于小肠上皮细胞的细胞质，主要位于小肠黏膜微绒毛的尖端。目前临床研究表明：I-FABP可以作为诊断早期肠黏膜损害和缺血的指标。而本实验通过测定化疗前后肿瘤患儿血浆中二胺氧化酶（DAO）及肠脂肪酸结合蛋白（I-FABP）的水平，同时相互比较其水平高低的差异，来研究不同强度化疗药物对肠粘膜屏障功能损伤的程度。 方法：将反复化疗的患儿按照化疗药物致吐强度的不同分为3组（轻度致吐组，中度致吐组,高度致吐组），采集各组患儿化疗前后不同时相（化疗前，化疗后第3天，化疗后第7天）的血浆，采用酶联免疫吸附测试法及分光光度计比色法，对血浆标本做DAO及I-FABP的水平测定；同时记录患儿化疗期间常规应用止吐药后仍出现的呕吐次数，根据呕吐次数多少分为强呕吐组（"g3次）及弱呕吐组（3次）；采集每日新鲜大便标本涂片革兰氏染色、高倍镜下读数，并计算出杆菌/细菌总数的比例。 计量资料采用均数±标准差(X±S)表示，计数资料采用相对比表示，实验数据用SPSS13.0软件进行统计学分析。检验水平α=0.05，P0.05视为差异有统计学意义。 结果： 1.不同致吐强度的各化疗组的DAO水平变化比较： （1）化疗前，不同致吐强度组之间的血浆DAO水平基本相同，各组间差异无统计学意义（P0.05）； （2）化疗后第3天及化疗后第7天，不同致吐强度组间的DAO水平存在显著性差异（P0.05），进一步分析发现，其中高致吐组DAO水平明显高于低、中致吐组，其差异有统计学意义（P0.05）； （3）化疗后第3天及化疗后第7天，各组血浆DAO水平均显著高于该组化疗前水平，各组前后差异均具有统计学意义（P0.05），进一步分析发现，化疗后第3天各组血浆DAO水平均高于该组化疗后第7天的DAO水平，各组差异均具有统计学意义（P0.05）。 2.不同致吐强度的各化疗组的I-FABP水平变化比较： （1）化疗前，不同致吐强度组之间的血浆I-FABP水平基本相同，各组间差异无统计学意义（P0.05）； （2）化疗后第3天及化疗后第7天，不同致吐强度组间的I-FABP水平存在显著性差异（P0.05），进一步分析发现，，高致吐组I-FABP水平明显高于低、中致吐组（P0.05），其差异均具有统计学意义（P0.05）； （3）化疗后第3天及化疗后第7天，各组血浆I-FABP水平均显著高于该组化疗前水平，各组前后差异具有统计学意义（P0.05），进一步分析发现，各组化疗后第3天血浆I-FABP水平均高于该组化疗后第7天血浆I-FABP水平，各组之间差异均具有统计学意义（P0.05）。 3.不同致吐强度的各化疗组之间的呕吐频率比较： （1）将各不同致吐强度组之间呕吐频率进行统计学分析，结果显示：高度组多于其他两组，而中度组多于低度组，但三组间差异无统计学意义（P0.05）。 （2）高度致吐组在化疗后第3天其呕吐频率达峰值，而中、低度组则于化疗后第4天呕吐频率达峰值。 4.呕吐频率与血浆DAO及I-FABP的关系： （1）直线相关分析结果显示，所有化疗患儿呕吐频率与血浆DAO水平呈直线正相关（r=0.744，P0.05）； （2）直线相关分析结果显示，所有化疗患儿呕吐频率与血浆I-FABP水平呈直线正相关（r=0.889，P0.05）； （3）在化疗第3天，强呕吐组的血浆DAO及I-FABP水平均显著高于弱呕吐组（P0.05）；在化疗第7天，强呕吐组的血浆DAO及I-FABP水平与弱呕吐组相比无明显差异（P0.05）。 5.不同致吐强度的各化疗组间的肠道菌群情况的比较： 对各致吐强度组间大便杆菌/细菌总数进行统计学分析发现，高度组低于其他两组，中度组低于低度组，但是三组之间差异无统计学意义（P0.05）。 6.肠道菌群与血浆DAO、I-FABP的关系： （1）直线相关分析结果显示，化疗组患儿的肠道菌群状况（杆菌/细菌总数比值）与血浆DAO水平呈负相关（r=-0.180，P0.05）； （2）直线相关分析结果显示，化疗组患儿的肠道菌群状况（杆菌/细菌总数比值）与血浆I-FABP水平呈负相关（r=-0.222，P0.05）。 结论： 1.各致吐组血浆DAO及I-FABP水平在化疗后第3天及第7天均显著高于该组化疗前水平，其中强呕吐组血浆DAO及I-FABP水平明显高于弱呕吐组，说明化疗患儿均存在不同程度胃肠黏膜屏障损伤，且随着化疗药物致吐强度的增加，胃肠粘膜损伤越严重，其引起的血浆DAO及I-FABP水平升高亦越明显。 2.化疗组患儿血浆DAO及I-FABP水平的变化与患儿呕吐频率均呈正相关，提示血浆DAO及I-FABP的变化可以反映患儿接受化疗后的胃肠道功能状态的改变情况。 3.化疗组患儿血浆DAO及I-FABP水平的变化与肠道菌群中杆菌/细菌总数呈负相关，提示化疗药物所导致的胃肠道损伤越重，肠道菌群失衡的表现就越明显。
[Abstract]:OBJECTIVE: Diamine oxidase (DAO) exists in mammalian mucosa or suprachorillary layer, most of which exist in intestinal mucosal villi, but few in endometrial villi; DAO is a highly active intracellular enzyme, its activity is related to the height of villi and the synthesis of nucleic acid and protein in mucosal cells. The changes of DAO content in tissues and plasma come from two pathways: one is that the necrotic intestinal mucosa cells fall off into the intestinal cavity, which leads to the decrease of DAO activity in the intestinal mucosa; the other is that the DAO in the intestinal cavity enters the lymphatic vessels and capillaries of the intestinal cell space. Therefore, in clinical and animal studies, we can reflect the function of intestinal mucosal barrier by measuring the level of DAO in blood and intestinal tissues, especially the activity of DAO in blood under non-invasive conditions to reflect intestinal injury and repair. Fatty acid binding protein (FABP) is a group of low molecular weight (about 15 kD) binding proteins with long-chain fatty acids. Its tissue specificity is strong. At present, it is believed that FABP exists in mammalian myocardium, small intestine, liver, adipose tissue, brain, epidermis and other specific tissues. It is a relatively new and valuable method to monitor tissue damage. Fatty acid binding protein (FABP) is one of the most important biological indicators. At present, the determination of FABP is mostly used in chronic heart failure (H-FABP), liver transplant rejection (L-FABP), non-heart-beating donor kidney viability selection (H-FABP, L-FABP), prevention of muscle injury (S-FABP) of professional athletes, and so on. TTY acid binding protein (I-FABP), one of the nine types of FABP, has high tissue specificity. It originates from the cytoplasm of small intestinal epithelial cells and is mainly located at the tip of small intestinal microvilli. The aim of this study was to determine the levels of diamine oxidase (DAO) and intestinal fatty acid binding protein (I-FABP) in plasma of children with cancer before and after chemotherapy, and to compare the differences between the levels of DAO and I-FABP.
Methods: The children with repeated chemotherapy were divided into three groups according to the intensity of vomiting induced by chemotherapeutic drugs (mild vomiting group, moderate vomiting group, high vomiting group). The plasma samples were collected at different phases before and after chemotherapy (before chemotherapy, 3 days after chemotherapy, 7 days after chemotherapy). Enzyme-linked immunosorbent assay (ELISA) and spectrophotometry were used to detect the plasma. The levels of DAO and I-FABP were measured, and the vomiting frequency was recorded after routine use of antiemetic drugs during chemotherapy. According to the vomiting frequency, the children were divided into strong vomiting group ("g3 times" and weak vomiting group ("3 times"); fresh stool samples were collected daily for Gram staining, high power microscopic reading, and the total number of bacilli/bacteria was calculated. Proportion.
The measurement data were expressed by mean [standard deviation] (X [S], the counting data were expressed by relative comparison, and the experimental data were statistically analyzed by SPSS 13.0 software.
Result:
1. the levels of DAO in different chemotherapeutic groups with different vomiting intensity were compared.
(1) Before chemotherapy, the levels of plasma DAO in different emetic intensity groups were basically the same, and there was no significant difference between each group (P 0.05).
(2) On the 3rd day after chemotherapy and the 7th day after chemotherapy, there were significant differences in the levels of DAO between the groups with different emetic intensity (P 0.05). Further analysis showed that the level of DAO in the high emetic group was significantly higher than that in the low emetic group, and the difference was statistically significant (P 0.05).
(3) On the 3rd day after chemotherapy and the 7th day after chemotherapy, the levels of plasma DAO in each group were significantly higher than those before chemotherapy, and the differences were statistically significant (P 0.05). Further analysis showed that the levels of plasma DAO in each group on the 3rd day after chemotherapy were higher than those on the 7th day after chemotherapy, and the differences were statistically significant (P 0.05).
2. the levels of I-FABP in different chemotherapeutic groups with different vomiting intensity were compared.
(1) Before chemotherapy, the plasma I-FABP levels of different emetic intensity groups were basically the same, and there was no significant difference between each group (P 0.05).
(2) On the third day after chemotherapy and the seventh day after chemotherapy, there was significant difference in the level of I-FABP between the groups with different emetic intensity (P 0.05). Further analysis showed that the level of I-FABP in the high emetic group was significantly higher than that in the low emetic group (P 0.05).
(3) On the 3rd day after chemotherapy and the 7th day after chemotherapy, the levels of plasma I-FABP in each group were significantly higher than those before chemotherapy, and the difference was statistically significant (P 0.05). Meaning (P0.05).
3. the frequency of vomiting between different chemotherapeutic groups with different vomiting intensity was compared:
(1) The vomiting frequency was statistically analyzed among different intensity groups. The results showed that the vomiting frequency in the high group was more than that in the other two groups, while the vomiting frequency in the middle group was more than that in the low group, but there was no significant difference among the three groups (P 0.05).
(2) The vomiting frequency peaked at the 3rd day after chemotherapy in the highly vomiting group, and peaked at the 4th day after chemotherapy in the middle and low vomiting group.
4. the relationship between the frequency of vomiting and plasma DAO and I-FABP:
(1) Linear correlation analysis showed that vomiting frequency was positively correlated with plasma DAO level (r = 0.744, P 0.05).
(2) Linear correlation analysis showed that vomiting frequency was positively correlated with plasma I-FABP level (r = 0.889, P 0.05).
(3) On the third day of chemotherapy, the plasma DAO and I-FABP levels in the strong vomiting group were significantly higher than those in the weak vomiting group (P 0.05); on the seventh day of chemotherapy, the plasma DAO and I-FABP levels in the strong vomiting group were not significantly different from those in the weak vomiting group (P 0.05).
5. comparison of intestinal microflora among different chemotherapeutic groups with different vomiting intensity:
Statistical analysis of the total fecal bacilli/bacteria counts among the vomiting intensity groups showed that the high group was lower than the other two groups, and the moderate group was lower than the low group, but there was no significant difference among the three groups (P 0.05).
6. the relationship between intestinal flora and plasma DAO and I-FABP:
(1) Linear correlation analysis showed that there was a negative correlation between the intestinal flora status (bacillus / bacteria ratio) and plasma DAO level (r = - 0.180, P 0.05).
(2) Linear correlation analysis showed that there was a negative correlation between the intestinal flora status (Bacillus/Bacterial count ratio) and plasma I-FABP level (r=-0.222, P 0.05).
Conclusion:
1. The plasma levels of DAO and I-FABP in each vomiting group were significantly higher than those before chemotherapy on the 3rd and 7th day after chemotherapy. The plasma levels of DAO and I-FABP in the strong vomiting group were significantly higher than those in the weak vomiting group, indicating that there were different degrees of gastrointestinal mucosal barrier damage in the chemotherapy group, and the more severe the gastrointestinal mucosal damage was with the increase of the intensity of vomiting induced by chemotherapy drugs. The level of plasma DAO and I-FABP increased significantly.
2. The changes of plasma DAO and I-FABP levels were positively correlated with the vomiting frequency, suggesting that the changes of plasma DAO and I-FABP could reflect the changes of gastrointestinal function after chemotherapy.
3. The changes of plasma DAO and I-FABP levels were negatively correlated with the total number of bacteria/bacteria in intestinal flora, suggesting that the more serious the gastrointestinal tract injury caused by chemotherapy drugs, the more obvious the imbalance of intestinal flora.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R730.5
本文编号：2229788
[Abstract]:OBJECTIVE: Diamine oxidase (DAO) exists in mammalian mucosa or suprachorillary layer, most of which exist in intestinal mucosal villi, but few in endometrial villi; DAO is a highly active intracellular enzyme, its activity is related to the height of villi and the synthesis of nucleic acid and protein in mucosal cells. The changes of DAO content in tissues and plasma come from two pathways: one is that the necrotic intestinal mucosa cells fall off into the intestinal cavity, which leads to the decrease of DAO activity in the intestinal mucosa; the other is that the DAO in the intestinal cavity enters the lymphatic vessels and capillaries of the intestinal cell space. Therefore, in clinical and animal studies, we can reflect the function of intestinal mucosal barrier by measuring the level of DAO in blood and intestinal tissues, especially the activity of DAO in blood under non-invasive conditions to reflect intestinal injury and repair. Fatty acid binding protein (FABP) is a group of low molecular weight (about 15 kD) binding proteins with long-chain fatty acids. Its tissue specificity is strong. At present, it is believed that FABP exists in mammalian myocardium, small intestine, liver, adipose tissue, brain, epidermis and other specific tissues. It is a relatively new and valuable method to monitor tissue damage. Fatty acid binding protein (FABP) is one of the most important biological indicators. At present, the determination of FABP is mostly used in chronic heart failure (H-FABP), liver transplant rejection (L-FABP), non-heart-beating donor kidney viability selection (H-FABP, L-FABP), prevention of muscle injury (S-FABP) of professional athletes, and so on. TTY acid binding protein (I-FABP), one of the nine types of FABP, has high tissue specificity. It originates from the cytoplasm of small intestinal epithelial cells and is mainly located at the tip of small intestinal microvilli. The aim of this study was to determine the levels of diamine oxidase (DAO) and intestinal fatty acid binding protein (I-FABP) in plasma of children with cancer before and after chemotherapy, and to compare the differences between the levels of DAO and I-FABP.
Methods: The children with repeated chemotherapy were divided into three groups according to the intensity of vomiting induced by chemotherapeutic drugs (mild vomiting group, moderate vomiting group, high vomiting group). The plasma samples were collected at different phases before and after chemotherapy (before chemotherapy, 3 days after chemotherapy, 7 days after chemotherapy). Enzyme-linked immunosorbent assay (ELISA) and spectrophotometry were used to detect the plasma. The levels of DAO and I-FABP were measured, and the vomiting frequency was recorded after routine use of antiemetic drugs during chemotherapy. According to the vomiting frequency, the children were divided into strong vomiting group ("g3 times" and weak vomiting group ("3 times"); fresh stool samples were collected daily for Gram staining, high power microscopic reading, and the total number of bacilli/bacteria was calculated. Proportion.
The measurement data were expressed by mean [standard deviation] (X [S], the counting data were expressed by relative comparison, and the experimental data were statistically analyzed by SPSS 13.0 software.
Result:
1. the levels of DAO in different chemotherapeutic groups with different vomiting intensity were compared.
(1) Before chemotherapy, the levels of plasma DAO in different emetic intensity groups were basically the same, and there was no significant difference between each group (P 0.05).
(2) On the 3rd day after chemotherapy and the 7th day after chemotherapy, there were significant differences in the levels of DAO between the groups with different emetic intensity (P 0.05). Further analysis showed that the level of DAO in the high emetic group was significantly higher than that in the low emetic group, and the difference was statistically significant (P 0.05).
(3) On the 3rd day after chemotherapy and the 7th day after chemotherapy, the levels of plasma DAO in each group were significantly higher than those before chemotherapy, and the differences were statistically significant (P 0.05). Further analysis showed that the levels of plasma DAO in each group on the 3rd day after chemotherapy were higher than those on the 7th day after chemotherapy, and the differences were statistically significant (P 0.05).
2. the levels of I-FABP in different chemotherapeutic groups with different vomiting intensity were compared.
(1) Before chemotherapy, the plasma I-FABP levels of different emetic intensity groups were basically the same, and there was no significant difference between each group (P 0.05).
(2) On the third day after chemotherapy and the seventh day after chemotherapy, there was significant difference in the level of I-FABP between the groups with different emetic intensity (P 0.05). Further analysis showed that the level of I-FABP in the high emetic group was significantly higher than that in the low emetic group (P 0.05).
(3) On the 3rd day after chemotherapy and the 7th day after chemotherapy, the levels of plasma I-FABP in each group were significantly higher than those before chemotherapy, and the difference was statistically significant (P 0.05). Meaning (P0.05).
3. the frequency of vomiting between different chemotherapeutic groups with different vomiting intensity was compared:
(1) The vomiting frequency was statistically analyzed among different intensity groups. The results showed that the vomiting frequency in the high group was more than that in the other two groups, while the vomiting frequency in the middle group was more than that in the low group, but there was no significant difference among the three groups (P 0.05).
(2) The vomiting frequency peaked at the 3rd day after chemotherapy in the highly vomiting group, and peaked at the 4th day after chemotherapy in the middle and low vomiting group.
4. the relationship between the frequency of vomiting and plasma DAO and I-FABP:
(1) Linear correlation analysis showed that vomiting frequency was positively correlated with plasma DAO level (r = 0.744, P 0.05).
(2) Linear correlation analysis showed that vomiting frequency was positively correlated with plasma I-FABP level (r = 0.889, P 0.05).
(3) On the third day of chemotherapy, the plasma DAO and I-FABP levels in the strong vomiting group were significantly higher than those in the weak vomiting group (P 0.05); on the seventh day of chemotherapy, the plasma DAO and I-FABP levels in the strong vomiting group were not significantly different from those in the weak vomiting group (P 0.05).
5. comparison of intestinal microflora among different chemotherapeutic groups with different vomiting intensity:
Statistical analysis of the total fecal bacilli/bacteria counts among the vomiting intensity groups showed that the high group was lower than the other two groups, and the moderate group was lower than the low group, but there was no significant difference among the three groups (P 0.05).
6. the relationship between intestinal flora and plasma DAO and I-FABP:
(1) Linear correlation analysis showed that there was a negative correlation between the intestinal flora status (bacillus / bacteria ratio) and plasma DAO level (r = - 0.180, P 0.05).
(2) Linear correlation analysis showed that there was a negative correlation between the intestinal flora status (Bacillus/Bacterial count ratio) and plasma I-FABP level (r=-0.222, P 0.05).
Conclusion:
1. The plasma levels of DAO and I-FABP in each vomiting group were significantly higher than those before chemotherapy on the 3rd and 7th day after chemotherapy. The plasma levels of DAO and I-FABP in the strong vomiting group were significantly higher than those in the weak vomiting group, indicating that there were different degrees of gastrointestinal mucosal barrier damage in the chemotherapy group, and the more severe the gastrointestinal mucosal damage was with the increase of the intensity of vomiting induced by chemotherapy drugs. The level of plasma DAO and I-FABP increased significantly.
2. The changes of plasma DAO and I-FABP levels were positively correlated with the vomiting frequency, suggesting that the changes of plasma DAO and I-FABP could reflect the changes of gastrointestinal function after chemotherapy.
3. The changes of plasma DAO and I-FABP levels were negatively correlated with the total number of bacteria/bacteria in intestinal flora, suggesting that the more serious the gastrointestinal tract injury caused by chemotherapy drugs, the more obvious the imbalance of intestinal flora.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R730.5
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